Co-Cultures of Oophila Amblystomatis Between Ambystoma Maculatum and Ambystoma Gracile Hosts Show Host-Symbiont Fidelity

A unique symbiosis occurs between embryos of the spotted salamander (Ambystoma maculatum) and a green alga (Oophila amblystomatis). Unlike most vertebrate host-symbiont relationships, which are ectosymbiotic, A. maculatum exhibits both an ecto- and an endo-symbiosis, where some of the green algal cells living inside egg capsules enter embryonic tissues as well as individual salamander cells. Past research has consistently categorized this symbiosis as a mutualism, making this the first example of a “beneficial” microbe entering vertebrate cells. Another closely related species of salamander, Ambystoma gracile, also harbors beneficial Oophila algae in its egg capsules. However, our sampling within the A. gracile range consistently shows this to be a strict ectosymbiotic interaction—with no sign of tissue or presumably cellular entry. In this study we swapped cultured algae derived from intracapsular fluid of different salamander hosts to test the fidelity of tissue entry in these symbioses. Both A. maculatum and A. gracile embryos were raised in cultures with their own algae or algae cultured from the other host. Under these in vitro culture conditions A. maculatum algae will enter embryonic A. maculatum tissues. Additionally, although at a much lower frequency, A. gracile derived algae will also enter A. maculatum host tissues. However, neither Oophila strain enters A. gracile hosts in these co-culture conditions. These data reveal a potential host-symbiont fidelity that allows the unique endosymbiosis to occur in A. maculatum, but not in A. gracile. However, preliminary trials in our study found that persistent endogenous A. maculatum algae, as opposed to the cultured algae used in subsequent trials, enters host tissues at a higher frequency. An analysis of previously published Oophila transcriptomes revealed dramatic differences in gene expression between cultured and intracapsular Oophila. These include a suite of genes in protein and cell wall synthesis, photosynthesis, central carbon metabolism suggesting the intracapsular algae are assimilating ammonia for nitrogen metabolism and may be undergoing a life-cycle transition. Further refinements of these co-culture conditions could help determine physiological differences between cultured and endogenous algae, as well as rate-limiting cues provided for the alga by the salamander